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Spliceosome Profiling Visualizes Operations of a Dynamic RNP at Nucleotide Resolution.


ABSTRACT: Tools to understand how the spliceosome functions in vivo have lagged behind advances in the structural biology of the spliceosome. Here, methods are described to globally profile spliceosome-bound pre-mRNA, intermediates, and spliced mRNA at nucleotide resolution. These tools are applied to three yeast species that span 600 million years of evolution. The sensitivity of the approach enables the detection of canonical and non-canonical events, including interrupted, recursive, and nested splicing. This application of statistical modeling uncovers independent roles for the size and position of the intron and the number of introns per transcript in substrate progression through the two catalytic stages. These include species-specific inputs suggestive of spliceosome-transcriptome coevolution. Further investigations reveal the ATP-dependent discard of numerous endogenous substrates after spliceosome assembly in vivo and connect this discard to intron retention, a form of splicing regulation. Spliceosome profiling is a quantitative, generalizable global technology used to investigate an RNP central to eukaryotic gene expression.

SUBMITTER: Burke JE 

PROVIDER: S-EPMC5940017 | biostudies-literature | 2018 May

REPOSITORIES: biostudies-literature

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Spliceosome Profiling Visualizes Operations of a Dynamic RNP at Nucleotide Resolution.

Burke Jordan E JE   Longhurst Adam D AD   Merkurjev Daria D   Sales-Lee Jade J   Rao Beiduo B   Moresco James J JJ   Yates John R JR   Li Jingyi Jessica JJ   Madhani Hiten D HD  

Cell 20180501 4


Tools to understand how the spliceosome functions in vivo have lagged behind advances in the structural biology of the spliceosome. Here, methods are described to globally profile spliceosome-bound pre-mRNA, intermediates, and spliced mRNA at nucleotide resolution. These tools are applied to three yeast species that span 600 million years of evolution. The sensitivity of the approach enables the detection of canonical and non-canonical events, including interrupted, recursive, and nested splicin  ...[more]

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